The objective of this program is to understand the regulation of the cell cycle dependent human histone genes. The histone genes are a multi-gene family of moderately repeated sequences with differences in the structure, organization and regulation of the various copies. The gene products, histone proteins, play a key role in the structural and functional properties of the eukaryotic genome. Histones package eukaryotic DNA into nucleosomes, and modifications in histone-DNA interactions are associated with changes in gene expression. The tight coupling of histone gene expression with DNA replication suggests a role for the histone genes in the control of cell growth and cell division. Studies carried out during the first two years of this program have clearly demonstrated that regulation of human histone gene expression includes both transcriptional and post-transcriptional components. These studies have also provided an initial indication of sequences involved in the regulation of two cell cycle dependent human histone genes. By deletion analysis and construction of chimeric genes that were assayed both in vitro and by transfection into mammalian cells, we have identified regions of these histone genes required for transcription, and shown that they undergo modifications in chromatin structure as a function of cell cycle dependent expression. Sequences within a histone mRNA that affect mRNA stability have been identified. We have also isolated from one of the histone gene clusters a fragment that can function as an origin of replication (ars element) in human cells. These results serve as the basis for our ongoing and proposed studies which are to: 1. Further define and characterize sequences that are required for 5' site specific initiation of transcription, that modulate levels of transcription, and that are responsible for cell cycle regulated expression of the human histone genes. 2. Characterize the modifications in histone gene chromatin structure that occur as a function of transcription during the cell cycle. 3. Isolate proteins that interact with specific regions of histone genes and examine their influence on chromatin structure and transcription. 4. Address the relationship between transcription and replication of human histone genes by further analysis of the structural and functional properties of the ars element we have isolated.